| Objective:Our objective was discuss the function of myoepithelial cell and epithelial cell in the progression of ductal carcinoma in situ to invasive ductal carcinoma Methods:In this study,we selected 157 cases of pure breast ductal carcinoma in situ in tianjin tumor hospital randomly.The all cases divided high grade,intermediate grade,low grade according to nucleus grade,the changes of myoepithelial cells in three groups of ductal carcinoma in situ,the changes of epithelial-mesenchymal transition(EMT)markers were stained with immunohistochemical staining,and carried out score and statistics;At the same time,163 cases of invasive ductal carcinoma that containing ductal carcinoma in situ were selected randomly,According to the expression state of ER,PR,HER-2,etc,the molecular typing group was divided.the immunohistochemical staining of EMT markers in each group,the expression difference between ductal carcinoma in situ and invasive ductal carcinoma was observed.According to immunohistochemical results,we implementation of cell experiments to verify the influence myoepithelial cell to glandular epithelia cell,selected myoepithelial cell line Hs 578 Bst,and glandular epithelial phenotype cell line MCF-7;Then,the co-culture system of Hs 578 Bst and glandular epithelial phenotype cell line MCF-7 was established.After 72 hours of contact with total of at37 ℃,the morphological changes of MCF-7 were examined by microscope.the functional changes of MCF-7,the expression level of EMT,and the EMT markers were measured by scratch test,transwell and western blot and QRT-PCR;The concentration of TGF-β1 in the co-culture fluid was detected by ELISA.Results:1、Immunohistochemical results showed that in situ carcinoma of the breast,the expression of myoepithelial cell was absent with the increaseing of nucleus grade.The expression of EMT related index(snail,zeb1)in glandular epithelial cellsincreased gradually with the increase nucleus grade,the expression of E-cad decreased gradually with the increase of nucleus grade,and there was obviously difference between the level of ductal carcinoma in situ(P < 0.05).In invasive ductal carcinoma(include DCIS),the expression of snail and zeb1 in invasive components were higher than in situ components,while the expression of E-cad in situ was lower than invasive components.Immunohistochemistry results prompt: the myoepithelial cell and glandular epithelial cells have changed.In our group,the previous experimental results showed that co-culture of myoepithelial and glandular epithelial which was man-made EMT,that the cell morphology and function of the myoepithelial changed was significantly,and the microenvironment and endocrine function of the myoepithelial changed was significantly,According to the above results,further study on the role of human epithelial cell to glandular epithelial from another direction,in this study,the glandular epithelial was added exogenous TGF-β1 as positive control,the co-culture model of myoepithelial and glandular epithelial cells was carried into the next step.the experimental results are as follows:2.MCF-7 cells and Hs 578 Bst cells cultured in vitro for 72 hours as experimental group,MCF-7 cells did not do any treatment as blank control,MCF-7added TGF-β1 as positive control,MCF-7 added TGF-β1 inhibitor as negative control.All these four groups cells were cultured at 37 ℃ for 72 hours.the results showed that the experimental group cells in the experimental group and the positive control group were similarly compared with the blank control group and the negative control group.Through scratch test,the value-added ability of each group cells,the results showed that the value-added ability of MCF-7 cells in experimental group and positive control group increased significantly.Transwell test showed that the invasive ability of the experimental group and positive control group was significantlyincreased than the blank group and the negative control group.The expression of EMT protein in each group was detected by western-blot test,the results showed that the expression of EMT protein in the experimental group and positive control group was higher,snail,twist,zeb1,E-cad and vimentin up-regulated,and the expression of E-cad decreased.RT PCR detected the gene expression level of correlation with EMT,and the results showed that: snail,twist,zeb1 and vimentin were up-regulatedin the experimental group and positive control group.The results of ELISA test showed that the concentration of TGF-β1 in the culture medium of co-culture was significantly increased compared with the control group(P < 0.05).Conclusion:In the process of invasion of breast ductal carcinoma,the expression of snail and zeb1 increased gradually with the increase of the level of ductal carcinoma in situ,the expression of E-cad gradually decreased gradually.Through the coculture of human myoepithelial cells and glandular epithelial cells,it can be found that the myoepithelial can effectively promote the invasion and migration of glandular epithelial cells.It shows that the myoepithelial promoted the glandular epithelial to occurrence EMT,and finally concludes that the myoepithelial and glandular epithelial cells interaction in the process of invasion in breast ductal carcinoma in situ. |
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